Changes in the function of transporters embedded into the cell membrane have been associated with erythrocyte dehydration in sickle cell anemia, Hb CC and SC diseases, hereditary spherocytosis and xerocytosis. Dehydrated cells are the most likely to sickle and produce vaso-occlusion;thus, specific therapies are being developed to diminish dehydration in vivo. The physiologically important regulators/determinants of activity of these pathways are not known, thus impeding the identification of subsets of patients more likely to experience cell dehydration and vaso-occlusion. We propose to identify physiologically important genes affecting the function of ion transport pathways and cell hydration in mouse and eventually human erythrocytes. We will use inbred mouse strains to identify quantitative tract loci (QTLs) and ultimately individual genes that modulate red cell hydration and transport. We have already identified inbred mouse strains with large differences in cell hydration: from the appropriate crosses, QTL analysis will be performed to focus on areas of the genome associated with the different phenotype, and identify the gene(s) involved. We also propose a detailed phenotypic characterization of the erythrocyte ion content and transport properties, focused on three pathways known to mediate dehydration (Gardos channel, K-CIcotransport, and Na-K pump), in 11 normal inbred mouse strains. These strains are widely used, with abundantly available genetic and phenotypic information. We will set-up informative crosses to identify specific QTLs associated with meaningful differences in transport activities. Definition of the genes influencing erythrocyte hydration and ion transport in the mouse will enlighten explorations in the sickle mouse model, and will allow testing of candidate genes using knock in/out experiments. The ultimate intent is to validate in future studies the mouse candidate genes with appropriate association studies in patients. We anticipate that these studies will lead to identification of novel genes relevant to pathophysiology of sickle cell syndromes and other anemias, and of potential new therapeutic targets, markers of disease severity, and indicators of susceptibility to specific therapies.